Androgen receptor antagonism drives cytochrome P450 17A1 inhibitor efficacy in prostate cancer
John D. Norris, … , William D. Figg, Donald P. McDonnell
John D. Norris, … , William D. Figg, Donald P. McDonnell
Published May 2, 2017
Citation Information: J Clin Invest. 2017;127(6):2326-2338. https://doi.org/10.1172/JCI87328.
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Research Article Endocrinology Therapeutics

Androgen receptor antagonism drives cytochrome P450 17A1 inhibitor efficacy in prostate cancer

Abstract

The clinical utility of inhibiting cytochrome P450 17A1 (CYP17), a cytochrome p450 enzyme that is required for the production of androgens, has been exemplified by the approval of abiraterone for the treatment of castration-resistant prostate cancer (CRPC). Recently, however, it has been reported that CYP17 inhibitors can interact directly with the androgen receptor (AR). A phase I study recently reported that seviteronel, a CYP17 lyase–selective inhibitor, ædemonstrated a sustained reduction in prostate-specific antigen in a patient with CRPC, and another study showed seviteronel’s direct effects on AR function. This suggested that seviteronel may have therapeutically relevant activities in addition to its ability to inhibit androgen production. Here, we have demonstrated that CYP17 inhibitors, with the exception of orteronel, can function as competitive AR antagonists. Conformational profiling revealed that the CYP17 inhibitor–bound AR adopted a conformation that resembled the unliganded AR (apo-AR), precluding nuclear localization and DNA binding. Further, we observed that seviteronel and abiraterone inhibited the growth of tumor xenografts expressing the clinically relevant mutation AR-F876L and that this activity could be attributed entirely to competitive AR antagonism. The results of this study suggest that the ability of CYP17 inhibitors to directly antagonize the AR may contribute to their clinical efficacy in CRPC.

Authors

John D. Norris, Stephanie J. Ellison, Jennifer G. Baker, David B. Stagg, Suzanne E. Wardell, Sunghee Park, Holly M. Alley, Robert M. Baldi, Alexander Yllanes, Kaitlyn J. Andreano, James P. Stice, Scott A. Lawrence, Joel R. Eisner, Douglas K. Price, William R. Moore, William D. Figg, Donald P. McDonnell

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Figure 1

CYP17 inhibitors bind and inhibit AR transcriptional activity.

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CYP17 inhibitors bind and inhibit AR transcriptional activity. (A) Struc...
(A) Structures of the compounds used in this study, including androgens (testosterone and R1881), benchmark antiandrogens (enzalutamide, bicalutamide, and hydroxyflutamide) and CYP17 inhibitors (seviteronel, galeterone, abiraterone, orteronel, and ketoconazole). (B) HEK293 cells were transfected with the WT-AR and treated with 0.1 nM [H3]-R1881 and a competitor ligand at the indicated doses for 2 hours. Lysates were subjected to scintillation counting and normalized to total protein amounts. Values in parentheses indicate the IC50 of each compound. Error bars represent the SD of duplicate samples from a representative experiment performed in triplicate. (C) CV1 cells were transfected with the WT-AR, MMTV-Luc, and Renilla-Luc and then treated with 0.1 nM R1881 and the indicated dose of antagonist for 24 hours. Dual-luciferase values were measured and normalized to Renilla-Luc. Error bars indicate the SD of triplicate samples of a representative experiment performed in triplicate. Quantitative PCR (qPCR) analysis was performed, and heatmaps were generated for AR target gene activation of LNCaP cells treated with (D) vehicle DMSO (Veh) or (E) 10 nM testosterone and 10 μM of the indicated ligand for 24 hours. (F) In-cell Western analysis using an AR antibody was performed on LNCaP cells treated with increasing concentrations of the indicated ligand for 24 hours. Geldanamycin (Geld), an HSP90 inhibitor, was included as a positive control for AR degradation. AR protein expression was assessed using the LI-COR Odyssey imaging system, normalizing to DRAQ5 (DNA dye). Error bars represent the SD of triplicate wells of a representative experiment performed in triplicate. (G) Western blot analysis was performed on LNCaP cells treated with vehicle DMSO, 1.0 nM testosterone (T), 1.0 μM geldanamycin, or 10 μM of antagonist for 24 hours. Blots were probed for the AR or β-actin as a loading control. Abi, abiraterone; Bic, bicalutamide; Enz, enzalutamide; Gal, galeterone; Ket, ketoconazole; OHF, hydroxyflutamide; Ort, orteronel; Sevi, seviteronel.